The coordinated events underlying protein folding, translocation across membranes, post-translational modifications and multi-unit assembly are essential to the viability of all cellular organisms. Recent data have demonstrated that conserved members of the "stress" or "heat shock" protein (HSP) family participate in these processing events by their ability to bind denatured or misfolded peptide sequences and then to release these polypeptide chains by an ATP-dependent mechanism. For a review of stress and heatshock proteins see Pelham, et al. Cell 46: 959-961, 1986, Hightower, et al., Cell 66: 191-197, 1991 and Gething, et al., Nature 355: 33-45, 1992. Although HSP members were first identified by their accumulation following cell exposure to elevated temperatures (Ritossa, et al. Experientia 18: 571-573, 1962; Tissieres et al., J. Mol. Biol. 84: 389-398, 1974), it was later recognized that a significant subset of these proteins was constitutively expressed and that cell incubations with a variety of metabolic poisons could result in gene induction (Welch et al., J. Biol. Chem. 258: 7102-7111, 1983). In eukaryotes, certain HSP products have been localized to specific cellular fractions, such as the cytosol, nucleus, endoplasmic reticulum or mitochondria, and recent experimental models have implicated these "chaperones" with facilitating protein transport across these specialized compartments (Chirico, et al. Nature 332: 805-810, 1988 and Deshaies et al., Nature 332: 800-805, 1988).
The stress70 gene comprise a group of related proteins within the larger HSP family. The stress70 gene family is complex and includes members from varied stages of evolution, including bacterial (dnaK, Bardwell, et al. Proc. Natl. Acad. Sci. 81: 848-852, 1984), yeast (kar2, ssa1, ssa2 and ssc1, Ingolia, et al. Mol. Cell. Biol. 2: 1388-1398, 1982; Chirico et al., supra; Deschaies et al., supra; Normington, et al. Cell 57: 1223-1236, 1989; and Rose, et al. Cell 57: 1211-1221, 1989) and mammalian species (hsp70, hsc70, grp78/BiP and pbp74, Mues, et al. J. Biol. Chem. 261: 874-877, 1986; Munro, et al. Cell 46: 291-300, 1986; Gething, et al. supra; and Domanico, et al. Mol. Cell. Biol. 13: 3598-3610, 1993). Structural analyses of these gene products have shown a highly conserved amino terminal domain that encodes ATP-binding and hydrolysis activity, and a less conserved carboxyl terminal portion that is required for peptide binding (Chappell, et al. J. Biol. Chem. 262: 746-751, 1987; Flajnik, et al. Immunogenetics 33: 295-300, 1991; and Rippmann, et al. EMBO J. 10: 1053-1059, 1991). The recognition that an hsp70-related gene encoded an abundant endoplasmic reticulum (ER) product identical to both the mammalian immunoglobulin binding protein (BiP) and the glucose-regulated protein (GRP78) suggested a specific role for stress70 proteins in the folding and assembly of newly synthesized proteins in the ER (Munro, et al. supra). This confirmed the hypothesis that stress70 molecules participate in protein processing during normal and stressed conditions (Bole, et al. J. Cell. Biol. 102: 1558-1566, 1986; Gething, et al. Cell 46: 939-950, 1986; and Pelham, et al. supra). GRP78/BiP has since been widely studied and shown to be targeted and retained in the ER by an amino terminal signal peptide and a carboxyl terminal ER retention signal (the tetrapeptide KDEL) (Munro, et al. Cell 48: 899-907, 1987 and Munro, et al., 1986 supra). Functional experiments have demonstrated that GRP78/BiP can reversibly bind in vitro to short peptides with relatively hydrophobic amino acid residues that are proposed to represent exposed domains on unfolded or misfolded proteins within the ER. In addition, GRP78/BiP encodes a peptide-stimulated ATPase activity that may then drive the protein folding process toward completion (Flynn, et al. Nature 353: 726-730, 1991 and Flynn, et al. Science 245: 385-390, 1989).
We describe the cloning of a constitutively expressed member of the stress70 protein chaperone family, designated Stch. Although Stch encodes a protein (STCH) with striking amino acid identity to HSP70 and BiP, it also has significant differences from the previously reported stress70 products. These unique differences include the presence of a unique hydrophobic signal sequence, a 50 amino acid insertion within the ATP-binding domain, and the absence of a carboxyl terminal peptide-binding domain. These structural features suggest the presence of a truncated stress70 product within protein secretory pathways that resembles the N-terminal proteolytic fragments of HSC70 and BiP (Chappell et al., 1987 supra and Kassenbrock, et al. EMBO J. 8: 1461-1467, 1989). Using antibodies to STCH antisera, we have confirmed these predictions and demonstrated that Stch encodes a smaller product (p60) which is highly enriched within the lumen of the cellular microsome fraction. In addition, we have shown data indicating that STCH expression varies between cell types. STCH exhibits ATPase activity that, in contrast to other HSP70-like molecules, is independent of peptide stimulation.